James T. McKenna, Stephen Thankachan, David S. Uygun, Charu Shukla, James M. McNally, Felipe L. Schiffino, Joshua Cordeira, Fumi Katsuki, Janneke C. Zant, Mackenzie C. Gamble, Karl Deisseroth, Robert W. McCarley, Ritchie E. Brown, Robert E. Strecker, Radhika Basheer

(A) Adeno-associated viral vectors with Cre-dependent expression of ArchT-green fluorescent proteins (AAV-Flex-ArchT-GFP) were bilaterally injected into the BF of PV-Cre mice. Following transduction, green (532 nm) laser light was applied bilaterally to excite ArchT and inhibit BF-PV neurons.

(B) Mice were placed in a recording chamber, where they were briefly exposed to increased CO2 or sound levels for 30 s every 5 min.

(C) AAV-ArchT-GFP injections transduced neurons and fibers throughout large areas of BF. One representative case is depicted here. Scale bar represents 1 mm. A summary of all cases is provided in .

(D) Representative primary traces of EEG, electromyography (EMG), laser on/off, and ambient CO2 percentage levels. Left: the gray shaded area shows arousal latency from the start of the stimulus to the point at which the animal awakens, determined as the onset of desynchronized EEG lasting >2 s, in response to increased CO2. Right: in the same mouse, the latency to arousal in response to increased CO2 was prolonged when BF-PV neurons were inhibited (CO2 w/ ArchT), as indicated by the wider gray shaded area.

(E) Mean latencies to arouse from CO2 (n = 7) significantly differed, comparing CO2 versus CO2 w/ ArchT (BF-PV inhibition).

(F) NREM probability curve plots indicate that the probability that animals stay in NREM sleep during CO2 infusion is significantly increased due to BF-PV optogenetic inhibition (CO2 w/ ArchT), compared to the laser off condition (CO2) or laser on, fluorophore-only controls (not shown; see main text).

(G) Representative primary traces of EEG, EMG, laser on/off, and ambient sounds levels. Left: the gray shaded area shows arousal latency from the start of the stimulus to the point at which the animal awakens, determined as the onset of desynchronized EEG lasting >2 s, in response to increased background sound levels. Right: in the same mouse, the latency to arousal in response to increased sound was prolonged when BF-PV neurons were inhibited (CO2 w/ ArchT), as indicated by the wider gray shaded area. Note: the first 10 s of sound stimuli were not detectable by the microphone above the background noise.

(H) Mean latencies to arouse in response to increased sound (n = 7) significantly differed, comparing sound versus sound w/ ArchT (BF-PV inhibition).

(I) NREM probability curve plots indicate that the probability that animals stay in NREM sleep during increased sound is significantly increased due to BF-PV optogenetic inhibition (sound w/ ArchT), compared to the laser off condition (Sound) or laser on, fluorophore-only controls (not shown; see main text).



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